Nearly 300 million individuals live with chronic hepatitis B virus (HBV) infection (CHB), for which no curative therapy is available. As viral diversity is associated with pathogenesis and immunological control of infection, improved methods to characterize this diversity could aid drug development efforts. Conventionally, viral sequencing data are mapped/aligned to a reference genome, and only the aligned sequences are retained for analysis. Thus, reference selection is critical, yet selecting the most representative reference a priori remains difficult. We investigate an alternative pangenome approach which can combine multiple reference sequences into a graph which can be used during alignment. Using simulated short-read sequencing data generated from publicly available HBV genomes and real sequencing data from an individual living with CHB, we demonstrate alignment to a phylogenetically representative \'genome graph\' can improve alignment, avoid issues of reference ambiguity, and facilitate the construction of sample-specific consensus sequences more genetically similar to the individual\'s infection. Graph-based methods can, therefore, improve efforts to characterize the genetics of viral pathogens, including HBV, and have broader implications in host-pathogen research.

Hepatitis B virus (HBV) genotype C is a prevalent HBV genotype in the Chinese population. Although genotype C shows higher sequence heterogeneity and more severe liver disease than other genotypes, its pathogenesis and immunological traits are not yet fully elucidated. In this study, we first established and chemically synthesized the consensus sequence based on representative 138 full-length HBV genotype C genomes from the Chinese population. The pHBV1.3C plasmid system, containing a 1.3-fold full-length HBV genotype C consensus sequence, was constructed for subsequent validation. Next, we performed functional assays to investigate the replicative competence of pHBV1.3C in vitro through the transient transfection of HepG2 and Huh7 cells and validated the in vivo function via a hydrodynamic injection to BALB/c recipient mice. The in vitro investigation revealed that the extracellular HBV DNA and intracellular replicative intermediate (i.e., pregenomic RNA, pgRNA) were apparently measurable at 48 h, and the HBsAg and HBcAg were still positive in hepatoma cells at 96 h. We also found that HBsAg and HBeAg accumulated at the extracellular and intracellular levels in a time-dependent manner. The in vivo validation demonstrated that pHBV1.3C plasmids induced HBV viremia, triggered morphological changes and HBsAg- or HBcAg- positivity of hepatocytes, and ultimately caused inflammatory infiltration and focal or piecemeal necrosis in the livers of the murine recipients. HBV protein (HBsAg) colocalized with CD8+ T cells or CD4+ T cells in the liver. F4/80+ Kupffer cells were abundantly recruited around the altered murine hepatocytes. Taken together, our results indicate that the synthetic consensus sequence of HBV genotype C is replication-competent in vitro and in vivo. This genotype C consensus genome supports the full HBV life cycle, which is conducive to studying its pathogenesis and immune response, screening novel antiviral agents, and further optimizing testing and therapeutics.

The International Pediatric Transplant Association convened an expert consensus conference to assess current evidence and develop recommendations for various aspects of care relating to post-transplant lymphoproliferative disorders after solid organ transplantation in children. In this report from the Viral Load and Biomarker Monitoring Working Group, we reviewed the existing literature regarding the role of Epstein-Barr viral load and other biomarkers in peripheral blood for predicting the development of PTLD, for PTLD diagnosis, and for monitoring of response to treatment. Key recommendations from the group highlighted the strong recommendation for use of the term EBV DNAemia instead of \"viremia\" to describe EBV DNA levels in peripheral blood as well as concerns with comparison of EBV DNAemia measurement results performed at different institutions even when tests are calibrated using the WHO international standard. The working group concluded that either whole blood or plasma could be used as matrices for EBV DNA measurement; optimal specimen type may be clinical context dependent. Whole blood testing has some advantages for surveillance to inform pre-emptive interventions while plasma testing may be preferred in the setting of clinical symptoms and treatment monitoring. However, EBV DNAemia testing alone was not recommended for PTLD diagnosis. Quantitative EBV DNAemia surveillance to identify patients at risk for PTLD and to inform pre-emptive interventions in patients who are EBV seronegative pre-transplant was recommended. In contrast, with the exception of intestinal transplant recipients or those with recent primary EBV infection prior to SOT, surveillance was not recommended in pediatric SOT recipients EBV seropositive pre-transplant. Implications of viral load kinetic parameters including peak load and viral set point on pre-emptive PTLD prevention monitoring algorithms were discussed. Use of additional markers, including measurements of EBV specific cell mediated immunity was discussed but not recommended though the importance of obtaining additional data from prospective multicenter studies was highlighted as a key research priority.

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Consensus PCR assays that can be used to sensitively detect several herpesvirus (HV) species across the different subfamilies were developed in this study. Primers containing degenerate bases were designed to amplify regions of the DNA polymerase (DPOL) gene of alpha- and gamma-HVs, and the glycoprotein B (gB) gene of beta-HVs in a singleplex, non-nested touchdown PCR format. The singleplex touchdown consensus PCR (STC-PCR) was used to amplify the DNA of eight human and 24 animal HVs. The assay was able to detect the lowest DNA dilution of 10-5 for alpha-HVs and 10-3 for beta- and gamma-HVs. In comparison, lowest detection limits of 10-5, 10-3, and 10-2 were obtained for alpha-, beta-, and gamma-HVs respectively when a nested PCR was used. The findings in this study suggest that the STC-PCR assays can be employed for the molecular surveys and clinical detection of novel and known HVs.

Cytomegalovirus (CMV) uveitis, a type of herpetic uveitis, is a major cause of infectious uveitis. Anterior and posterior CMV uveitis have diverse clinical presentations and treatment modalities. Based on expert consensus in Taiwan, this article provides suggestions regarding clinical manifestations, diagnosis, and treatment strategies for CMV uveitis based on clinical practice experience in Taiwan. CMV uveitis may have a distinct clinical presentation. Polymerase chain reaction (PCR) is an essential diagnostic tool to confirm a diagnosis. Antiviral therapy is the mainstay of treatment. Different agents, routes, and other supplemental treatments have been summarized and discussed in this article. Early diagnosis and appropriate treatment of CMV uveitis are crucial to avoid irreversible complications and vision loss. This consensus provides practical guidelines for ophthalmologists in Taiwan.

UNASSIGNED: Nucleos(t)ide analogues (NAs) are effective in suppressing the replication of the hepatitis B virus. However, NAs cannot effectively induce hepatitis B surface antigen (HBsAg) seroclearance, which represents the optimal treatment endpoint in chronic hepatitis B (CHB). Hence, most CHB patients are advised for indefinite NA therapy, but recent data has supported the concept of finite NA therapy before HBsAg seroclearance.
UNASSIGNED: This article covered the latest evidence on stopping NAs in CHB, with a focused analysis on international guidelines. Articles were retrieved by a literature search on PubMed with the keywords \'chronic hepatitis B,\' \'antiviral therapy,\' \'nucleos(t)ide analogue,\' \'cessation,\' \'stopping\', and \'finite.\' Studies up till 1 December 2022 were included.
UNASSIGNED: Finite NA therapy in CHB has the potential in enhancing HBsAg seroclearance, however it also carries rare but potentially severe risks. NA cessation before HBsAg seroclearance is only suitable for a highly selected group of patients, whereas the majority of CHB patients should be treated indefinitely or until HBsAg seroclearance. Current guidelines have provided recommendations on stopping NAs, but further research is required to optimize the monitoring and retreatment protocol after stopping NAs.

A major goal of curative hepatitis B virus (HBV) treatments is the reduction or inactivation of intrahepatic viral covalently closed circular DNA (cccDNA). Hence, precise cccDNA quantification is essential in preclinical and clinical studies. Southern blot (SB) permits cccDNA visualisation but lacks sensitivity and is very laborious. Quantitative PCR (qPCR) has no such limitations but inaccurate quantification due to codetection of viral replicative intermediates (RI) can occur. The use of different samples, preservation conditions, DNA extraction, nuclease digestion methods and qPCR strategies has hindered standardisation. Within the ICE-HBV consortium, available and novel protocols for cccDNA isolation and qPCR quantification in liver tissues and cell cultures were compared in six laboratories to develop evidence-based guidance for best practices.
Reference material (HBV-infected humanised mouse livers and HepG2-NTCP cells) was exchanged for cross-validation. Each group compared different DNA extraction methods (Hirt extraction, total DNA extraction with or without proteinase K treatment (+PK/-PK)) and nuclease digestion protocols (plasmid-safe ATP-dependent DNase (PSD), T5 exonuclease, exonucleases I/III). Samples were analysed by qPCR and SB.
Hirt and -PK extraction reduced coexisting RI forms. However, both cccDNA and the protein-free relaxed circular HBV DNA (pf-rcDNA) form were detected by qPCR. T5 and Exo I/III nucleases efficiently removed all RI forms. In contrast, PSD did not digest pf-rcDNA, but was less prone to induce cccDNA overdigestion. In stabilised tissues (eg, Allprotect), nucleases had detrimental effects on cccDNA.
We present here a comprehensive evidence-based guidance for optimising, controlling and validating cccDNA measurements using available qPCR assays.

Viral corneal endotheliitis, a blinding corneal disease, is often misdiagnosed due to the diverse clinical manifestations, complex conditions, unclear diagnostic criteria, and limited methods of ocular virus detection. In addition, there is still a lack of unified and standardized treatment for viral corneal endotheliitis. The consensus, basing upon the latest research progress and expert recommendations regarding the clinical care of patients with viral corneal endotheliitis, targets to offer best practice advice for the clinical management of viral corneal endotheliitis and has been fully discussed by the experts of the Ocular Infection Group of Chinese Ophthalmologist Association.
病毒性角膜内皮炎是一种致盲性角膜病，因临床表现多样，病情复杂，诊断标准不明确以及眼部病毒检测手段有限等原因，易误诊或漏诊，且目前尚缺乏统一和规范的治疗方案。鉴于此，中国医师协会眼科医师分会眼感染学组汇集国内眼感染和角膜病专家，以国内外相关最新研究为基础，参考国内相关专家的临床经验，经过充分讨论，针对病毒性角膜内皮炎的临床诊疗形成共识性意见，以期为临床开展工作提供参考和指导。.

UNASSIGNED: Epidemiologic data regarding chronic hepatitis B virus infections in Israel is limited as extensive population-based studies have not been performed.
UNASSIGNED: This work aimed to evaluate the current characteristics of hepatitis B infection among Israeli adults and evaluate adherence to the European Association for the Study of the Liver practice guidelines for antiviral treatment.
UNASSIGNED: Clinical and demographic data of HBsAg-positive patients registered in the Leumit-Health-Service database (one of the four major health maintenance organizations in Israel) between 2000 and 2019 were retrieved. Patients were compared according to eligibility to antiviral treatment and type of nucleos(t)ide analogue (NA) treatment.
UNASSIGNED: In total, 1216 patients had documented HBsAg positivity (males 58.6%, mean age 40.2 ± 14.2 years), 90.6% of whom were HBeAg negative. Antiviral therapy eligibility was met by 37% of patients, among whom 89% received antiviral therapy. Antiviral therapies include NA with a high barrier to resistance (HBR) (64.5%) and NA with a low barrier to resistance (LBR) (35.5%). Compared to patients who received LBR NA, patients receiving HBR NA had shorter treatment (68.7 ± 50 vs. 161.5 ± 42.6 months, p < .001) and follow-up duration (125 ± 68 vs. 188 ± 48 months, p < .001); at the end of follow-up, ALT levels and APRI score were higher among patients on LBR NA compared to patients on HBR NA.
UNASSIGNED: Most patients received antiviral treatment according to the international practice guidelines. However, one-third of them were treated with a less potent NA, probably due to their lower cost. These findings should encourage the optimization of HBV care and full compliance with the professional practice guideline recommendations.